Abstract: Medical treatment with the aminoglycosidic antibiotic gentamicin may produce side effects that include neuromuscular blockage and ototoxicity; which are believed to result from a dysfunction of nicotinic acetylcholine receptors (AChRs). Gentamicin is known to reversibly block ACh-currents generated by the activation of muscle-type alphabetagammadelta-AChR and neuronal alpha9-AChR. We studied the effects of gentamicin on heteromeric alphabetagammadelta-AChR and homomeric alpha7-AChR expressed in Xenopus oocytes. Prolonged treatment with gentamicin, and other antibiotics, differentially altered alphabetagammadelta- and alpha7-AChR responses. Specifically, gentamicin accelerated desensitization and did not reduce ACh-currents in oocytes expressing alphabetagammadelta-AChRs, whereas ACh-currents were reduced and desensitization was unaltered in oocytes expressing alpha7-AChRs. Moreover, acutely applied gentamicin acted as a competitive antagonist on both types of receptors and increased the rate of desensitization in alphabetagammadelta-AChR while reducing the rate of desensitization in alpha7-AChR. This data helps to better understand the action of gentamicin on muscle and nervous tissues, providing mechanistic insights that could eventually lead to improving the medical use of aminoglycosides.

Abstract: Nicotinic acetylcholine receptors (AChRs) are one of the best characterized ion channels from the Cys-loop receptor superfamily. The study of acetylcholine binding proteins and prokaryotic ion channels from different species has been paramount for the understanding of the structure-function relationship of the Cys-loop receptor superfamily. AChR function can be modulated by different ligand types. The neurotransmitter ACh and other agonists trigger conformational changes in the receptor, finally opening the intrinsic cation channel. The so-called gating process couples ligand binding, located at the extracellular portion, to the opening of the ion channel, located at the transmembrane region. After agonist activation, in the prolonged presence of agonists, the AChR becomes desensitized. Competitive antagonists overlap the agonist-binding sites inhibiting the pharmacological action of agonists. Positive allosteric modulators (PAMs) do not bind to the orthostetic binding sites but allosterically enhance the activity elicited by agonists by increasing the gating process (type I) and/or by decreasing desensitization (type II). Instead, negative allosteric modulators (NAMs) produce the opposite effects. Interestingly, this negative effect is similar to that found for another class of allosteric drugs, that is, noncompetitive antagonists (NCAs). However, the main difference between both categories of drugs is based on their distinct binding site locations. Although both NAMs and NCAs do not bind to the agonist sites, NACs bind to sites located in the ion channel, whereas NAMs bind to nonluminal sites. However, this classification is less clear for NAMs interacting at the extracellular-transmembrane interface where the ion channel mouth might be involved. Interestingly, PAMs and NAMs might be developed as potential medications for the treatment of several diseases involving AChRs, including dementia-, skin-, and immunological-related diseases, drug addiction, and cancer. More exciting is the potential combination of specific agonists with specific PAMs. However, we are still in the beginning of understanding how these compounds act and how these drugs can be used therapeutically.

Abstract: Tubby and tubby-like proteins (TULPs) are encoded by members of a small gene family. An autosomal recessive mutation in the mouse tub gene leads to blindness, deafness, and maturity-onset obesity. The mechanisms by which the mutation causes the obesity syndrome has not been established. We compared obese tub/tub mice and their lean littermates in order to find abnormalities within the mediobasal hypothalamus, a region intimately associated with the regulation of body weight. Using an antiserum to the vesicular acetylcholine transporter (VAChT), a marker for cholinergic neurons, many unusually large VAChT-immunoreactive (-ir) nerve terminals, identified by colocalization with the synaptic vesicle protein synaptophysin, were demonstrated in the hypothalamic arcuate nucleus of obese tub/tub mice. Double-labeling showed that VAChT-ir nerve endings also contained glutamic acid decarboxylase (GAD), a marker for gamma-aminobutyric acid (GABA) neurons. The VAChT- and GAD-ir nerve terminals were in close contact with blood vessels, identified with antisera to platelet endothelial cell adhesion molecule-1 (PECAM; also called CD31), laminin, smooth muscle actin (SMA), and glucose transporter-1 (GLUT1). Such large cholinergic and GABAergic nerve terminals surrounding blood vessels were not seen in the arcuate nucleus of lean tub/+ mice. The presence of abnormal cholinergic/GABAergic vascular innervation in the arcuate nucleus suggests that alterations in this region, which contains neurons that receive information from the periphery and which relays information about the energy status to other parts of the brain, may be central in the development of the obese phenotype in animals with an autosomal recessive mutation in the tub gene.

Abstract: Functional foods and nutraceuticals are becoming popular alternatives to pharmacological treatments by providing health benefits and (or) reducing the risk of chronic diseases. Flaxseed is a rich source of 3 components with demonstrated cardioprotective effects: the omega-3 fatty acid alpha-linolenic acid (ALA), dietary fibre, and phytoestrogen lignans. Multiple clinical dietary intervention trials report that consuming flaxseed daily can modestly reduce circulating total cholesterol (TC) by 6%-11% and low-density lipoprotein (LDL) cholesterol by 9%-18% in normolipemic humans and by 5%-17% for TC and 4%-10% for LDL cholesterol in hypercholesterolemic patients, as well as lower various markers associated with atherosclerotic cardiovascular disease in humans. Evidence to date suggests that the dietary fibre and (or) lignan content of flaxseed provides the hypocholesterolemic action. The omega-3 ALA found in the flaxseed oil fraction also contributes to the antiatherogenic effects of flaxseed via anti-inflammatory and antiproliferative mechanisms. Dietary flaxseed may also protect against ischemic heart disease by improving vascular relaxation responses and by inhibiting the incidence of ventricular fibrillation.

Abstract: The intestinal microbiota is a diverse and dynamic ecosystem,1 which has developed a mutualistic relationship with its host and plays a crucial role in the development of the host's innate and adaptive immune responses.2 This ecosystem serves the host by protecting against pathogens, harvesting otherwise inaccessible nutrients, aiding in neutralisation of drugs and carcinogens, and affecting the metabolism of lipids.3 Gut bacteria modulate intestinal motility, barrier function and visceral perception.4
An interaction between the intestinal microbiota and the central nervous system (CNS) may seem difficult to conceive at first sight, but clinicians are well aware of the benefit of oral antibiotics and laxatives in the treatment of hepatic encephalopathy.5 Data accumulated from animal studies indicate that there is central sensing of gastrointestinal infections. For example, acute infection with Campylobacter jejuni results in anxiety-like behaviour and rapid activation of vagal pathways prior to onset of immune responses,6 while chronic Helicobacter pylori infection in mice leads to abnormal feeding behaviour and upregulation of tumour necrosis factor &#945; (TNF&#945;) in the median eminence of the hypothalamus.7 Rapid and sustained gut&#65533;brain communication may confer a significant advantage to the host, as central activation in response to changes in commensals or pathogens would allow better control of gut function and immunity.

Abstract: Isoquinoline alkaloids represent a group of natural products with remarkable importance in the contemporary biomedical research and drug discovery programs. Several members of this group exhibit immense pharmacological and biological properties, including potential anticancer properties. Although the molecular targets of these alkaloids are not yet clearly delineated, extensive research in this area continues to build up new data that are clinically exploitable. The gross structural features of many of the members DNA interaction are more or less clear, but the mystery still remains on many aspects of their binding, including specificity and energetics. RNA-binding aspects of these alkaloids are being elucidated. More recent advancements in analytical instrumentation have enabled clearer elucidation and correlation of the structural and energetic aspects of the interaction. In this review, we report up-to-date details of the interaction of berberine, palmatine, and jatrorrhizine of the protoberberine group, sanguinarine from the benzophananthridine group, and several of their synthetic derivatives, such as coralyne, berberrubine, palmatrubine, and jatrorubin with nucleic acids have been reviewed. These studies, taken together up to now, have led to interesting knowledge on the mode, mechanism, specificity of binding, and correlation between structural aspects and energetics enabling a complete set of guidelines for design of new drugs. In contemporary research, several derivatives of these natural alkaloids are being prepared and investigated in several laboratories for ultimate discovery of new compounds that can be used as effective therapeutic agents.

Abstract: The medial prefrontal cortex (mPFC) has been implicated in modulating anxiety. However, it is unknown whether excitatory or inhibitory neurotransmission in the infralimbic (IL) subregion of the mPFC underlies the pathology of anxiety-related behavior. To address this issue, we infused the GABAA receptor (GABAAR) antagonist bicuculline to temporarily activate the IL cortex. IL cortex activation decreased the time spent in the center area in the open field test, decreased exploration of the open-arms in the elevated plus maze test, and increased the latency to bite food in the novelty-suppressed feeding test. These findings substantiate the GABAergic system's role in anxiety-related behaviors. IL cortex inactivation with the AMPA receptor (AMPAR) antagonist CNQX produced opposite, anxiolytic effects. However, infusion of the NMDA receptor (NMDAR) antagonist AP5 into the IL cortex had no significant effect. Additionally, we did not observe motor activity deficits or appetite deficits following inhibition of GABAergic or glutamatergic neurotransmission. Interestingly, we found parallel and corresponding electrophysiological changes in anxious mice; compared to mice with relatively low anxiety, the relatively high anxiety mice exhibited smaller evoked inhibitory postsynaptic currents (eIPSCs) and larger AMPA-mediated evoked excitatory postsynaptic currents (eEPSCs) in pyramidal neurons in the IL cortex. The changes of eIPSCs and eEPSCs were due to presynaptic mechanisms. Our results suggest that imbalances of neurotransmission in the IL cortex may cause a net increase in excitatory inputs onto pyramidal neurons, which may underlie the pathogenic mechanism of anxiety disorders.

Abstract: We embarked on a study to delineate the behavioural changes in mice after 4 weeks of voluntary exercise. As an initial behavioural characterization, we exposed the control and exercising mice to a modified hole board and an open field test. As compared to control mice, exercising animals showed clear signs of increased behavioural inhibition (e.g. a longer latency to enter unprotected areas), suggesting increased anxiety in these animals. In addition, the exercising mice were reluctant to spend time in the open field's centre during the beginning of the 30-min open field test, but compensated for this at later times. Paradoxically, the exercising animals showed more rearings on the board of the modified hole board, indicating decreased anxiety. Thus, the behavioural inhibition seen in exercising mice is likely to represent decreased stress responsiveness at the behavioural level which can also be interpreted as reduced impulsiveness. To clarify whether voluntary exercise evolves in more or less anxiety-related behaviour, we exposed animals to the elevated plus-maze and the dark-light box, two selective tests for unconditioned anxiety. Clearly, compared to the control animals, exercising mice spent significantly more time on the open arm of the plus-maze and spent double the amount of time in the light compartment of the dark-light box. Taken together, we conclude that long-term voluntary exercise appears to result in decreased anxiety-related behaviour and impulsiveness. Thus, our observations fit into the concept that regular exercise strengthens endogenous stress coping mechanisms, thereby protecting the organism against the deleterious effects of stress.

Abstract: BACKGROUND AND PURPOSE: Several agonists of the alpha7 nicotinic acetylcholine receptor (nAChR) have been developed for treatment of cognitive deficits. However, agonist efficacy in vivo is difficult to reconcile with rapid alpha7 nAChR desensitization in vitro; and furthermore, the correlation between in vitro receptor efficacy and in vivo behavioural efficacy is not well delineated. The possibility that agonists of this receptor actually function in vivo as inhibitors via desensitization has not been finally resolved. EXPERIMENTAL APPROACH: Two structurally related alpha7 nAChR agonists were characterized and used to assess the degree of efficacy required in a behavioural paradigm. KEY RESULTS: NS6784 activated human and rat alpha7 nAChR with EC(50)s of 0.72 and 0.88 microM, and apparent efficacies of 77 and 97% respectively. NS6740, in contrast, displayed little efficacy at alpha7 nAChR (<2% in oocytes, < or =8% in GH4C1 cells), although its agonist-like properties were revealed by adding a positive allosteric modulator of alpha7 nAChRs or using the slowly desensitizing alpha7V274T receptor. In mouse inhibitory avoidance (IA) memory retention, NS6784 enhanced performance as did the 60% partial agonist A-582941. In contrast, NS6740 did not enhance performance, but blocked effects of A-582941. CONCLUSIONS AND IMPLICATIONS: Collectively, these findings suggest that a degree of alpha7 nAChR agonist efficacy is required for behavioural effects in the IA paradigm, and that such behavioural efficacy is not due to alpha7 nAChR desensitization. Also, a partial agonist of very low efficacy for this receptor could be used as an inhibitor, in the absence of alpha7 nAChR antagonists with favourable CNS penetration.

Abstract: The lipid chaperone aP2/FABP4 has been implicated in the pathology of many immunometabolic diseases, including diabetes in humans, but aP2 has not yet been targeted for therapeutic applications. aP2 is not only an intracellular protein but also an active adipokine that contributes to hyperglycemia by promoting hepatic gluconeogenesis and interfering with peripheral insulin action. Serum aP2 levels are markedly elevated in mouse and human obesity and strongly correlate with metabolic complications. These observations raise the possibility of a new strategy to treat metabolic disease by targeting serum aP2 with a monoclonal antibody (mAb) to aP2. We evaluated mAbs to aP2 and identified one, CA33, that lowered fasting blood glucose, improved systemic glucose metabolism, increased systemic insulin sensitivity, and reduced fat mass and liver steatosis in obese mouse models. We examined the structure of the aP2-CA33 complex and resolved the target epitope by crystallographic studies in comparison to another mAb that lacked efficacy in vivo. In hyperinsulinemic-euglycemic clamp studies, we found that the antidiabetic effect of CA33 was predominantly linked to the regulation of hepatic glucose output and peripheral glucose utilization. The antibody had no effect in aP2-deficient mice, demonstrating its target specificity. We conclude that an aP2 mAb-mediated therapeutic constitutes a feasible approach for the treatment of diabetes.